Project Summary Although much remains unclear with respect to the pathogenesis of COVID-19, the possibilities that cytokine storm and altered coagulation contribute to adverse disease pathogenesis have emerged. Both the cytokine output and promotion of coagulation may be mediated by activated monocytes. Monocytes, for instance, are the central leukocyte in blood to express tissue factor (F3) and initiate coagulation and this activity promotes comorbidity in Ebola and HIV infections. One cytokine that seems most notably elevated in plasma of COVID patients is IL-6. In keeping with the potential importance of IL-6 in cytokine storm, our preliminary data reveal that blood monocytes, of all subsets, are a major source of IL-6 in COVID leukocytes and that their production of IL-6 positively correlates with adverse disease progression. Surprisingly, though monocytes produce cytokines, other features of canonical activation were not present in monocyte subsets of COVID patients. Particularly, induction of functional tissue factor in IL-6-producing monocytes was minimal to absent. This was in striking contrast to control monocytes from healthy subjects treated ex vivo with LPS or resiquimod. Given the clinical concern that coagulation may be altered in COVID-19 and contribute to pathogenesis of adverse disease and given the very robust expression of IL-6, we were especially surprised that tissue factor was not induced. Overall, it appears that the stimulus for activation of monocytes in COVID-19 patients is distinct from canonical responses that also induce tissue factor in cytokine-producing cells, or perhaps a subset of monocytes able to activate the tissue factor pathway is missing or in extracted COVID-19 PBMCs. The overarching aim of this work is to define, using a robust longitudinal dataset, whether proinflammatory activation of monocyte subsets in blood associates with disease severity and to define the core characteristics of such activation. We will also carry out exploratory analyses in search of signals that lead to such activation. A key resource for our proposal is access to a bank of frozen PBMC, serum, plasma and whole blood in which longitudinal blood draws are being collected on 300 COVID-19 patients of differing disease severity admitted to Barnes Jewish Hospital. This bank has been established by the Institutional Clinical and Translational Research program at Washington University School of Medicine. Resources from this bank will be coupled with a stock of frozen PBMCs from >50 control participants in our laboratory and PBMCs from HIV subjects, where the state of monocyte activation will be compared with that of monocytes derived from COVID patients.